NAD(P)H quinone oxidoreductase‐1 (NQO1) is a homodimeric protein that acts as a detoxifying enzyme or as a chaperone protein. Dicourmarol interacts with NQO1 at the NAD(P)H binding site and can both inhibit enzyme activity and modulate the interaction of NQO1 with other proteins. We show that the binding of dicoumarol and related compounds to NQO1 generates negative cooperativity between the monomers. This does not occur in the presence of the reducing cofactor, NAD(P)H, alone. Alteration of Gly150 (but not Gly149 or Gly174) abolished the dicoumarol‐induced negative cooperativity. Analysis of the dynamics of NQO1 with the Gaussian network model indicates a high degree of collective motion by monomers and domains within NQO1. Ligand binding is predicted to alter NQO1 dynamics both proximal to the ligand binding site and remotely, close to the second binding site. Thus, drug‐induced modulation of protein motion might contribute to the biological effects of putative inhibitors of NQO1.

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HighlightsSNP rs1143684 results in either a Phe or Leu at position 47 in human NQO2.NQO2-L47 has a slightly lower kcat/Km; it also has a lower Ki,app with resveratrol.NQO2-L47 is more unstable to proteolysis and thermal denaturation.NQO2-F47 (but not NQO2-L47) shows negative cooperativity towards resveratrol.NQO2-L47 exists as multiple conformers in solution.

Osteoporosis is a common disease characterised by reduced bone mass and an increased risk of fragility fractures. Low bone mineral density is known to significantly increase the risk of osteoporotic fractures, however, the majority of non-traumatic fractures occur in individuals with a bone mineral density too high to be classified as osteoporotic. Therefore, there is an urgent need to investigate aspects of bone health, other than bone mass, that can predict the risk of fracture. Here, we successfully predicted association between bone collagen and nail keratin in relation to bone loss due to oestrogen deficiency using Raman spectroscopy. Raman signal signature successfully discriminated between ovariectomised rats and their sham controls with a high degree of accuracy for the bone (sensitivity 89%, specificity 91%) and claw tissue (sensitivity 89%, specificity 82%). When tested in an independent set of claw samples the classifier gave 92% sensitivity and 85% specificity. Comparison of the spectral changes occurring in the bone tissue with the changes occurring in the keratin showed a number of common features that could be attributed to common changes in the structure of bone collagen and claw keratin. This study established that systemic oestrogen deficiency mediates parallel structural changes in both the claw (primarily keratin) and bone proteins (primarily collagen). This strengthens the hypothesis that nail keratin can act as a surrogate marker of bone protein status where systemic processes induce changes.

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The National Cancer Institute chemical database has been screened using in silico docking to identify novel nanomolar inhibitors of NRH:quinone oxidoreductase 2 (NQO2). The inhibitors identified from the screen exhibit a diverse range of scaffolds and the structure of one of the inhibitors, NSC13000 cocrystalized with NQO2, has been solved. This has been used to aid the generation of a structure-activity relationship between the computationally derived binding affinity and experimentally measured enzyme inhibitory potency. Many of the compounds are functionally active as inhibitors of NQO2 in cells at nontoxic concentrations. To show this, advantage was taken of the NQO2-mediated toxicity of the chemotherapeutic drug CB1954. The toxicity of this drug is substantially reduced when the function of NQO2 is inhibited, and many of the compounds achieve this in cells at nanomolar concentrations. The NQO2 inhibitors also attenuated TNFa-mediated, NF-kB-driven transcriptional activity. The link between NQO2 and the regulation of NF-kB was confirmed by using short interfering RNA to NQO2 and by the observation that NRH, the cofactor for NQO2 enzyme activity, could regulate NF-kB activity in an NQO2-dependent manner. NF-kB is a potential therapeutic target and this study reveals an underlying mechanism that may be usable for developing new anticancer drugs.

No abstract

Patients diagnosed with osteoporosis have reported loss of fingernail resilience as the disease progresses. Keratin is the predominant protein in human nail tissue, and its structure has been postulated to be different in fingernails clipped from subjects who have sustained fragility fractures and those who have not, which may offer a window into the donor's bone health. This study was designed to qualify these differences, which may lead to the development of a novel screening tool for fracture risk. Raman spectroscopy was used to measure the fingernails of 633 postmenopausal women who presented at six fracture clinics located across the UK and Ireland. The Raman signals from donor's fingernails were compared between (1) fracture and nonfracture and (2) osteoporotic versus non‐osteoporotic donors The data presented show differences in the protein changes observed for pervasive osteoporosis compared to a general increased risk of fragility fracture. For fracture risk, compositional changes falling into broad classes of amino acid residue (aliphatic, aromatic, acidic, amide and sulphurous) were observed, while a difference in disulphide bonding levels was reaffirmed. For pervasive osteoporosis, the disulphide mode suggested increasing disorder in disulphide bonding orientation. Fractures were associated with a transition from alpha helical secondary structure to random, while the pervasive osteoporosis cases were associated with a transition to beta sheet structure. General fracture risk is associated with a change in the structure and composition of the keratin protein. Osteoporosis is associated with different protein structural changes and an increase in free acid groups. Copyright © 2017 John Wiley & Sons, Ltd.